Human African trypanosomiasis (HAT) is caused by the parasitic protozoan, Trypanosoma brucei. HAT is listed as a WHO Category 1 disease (emerging and uncontrolled) that exerts a large burden in both health and economic costs to the endemic regions in Africa. The disease is fatal unless treated and current therapies suffer from high toxicity and difficult treatment regimes. Furthermore none of the current drugs are effective against both species and stages of the disease. There is a great need to translate recent advances in the understanding of the basic biology of the parasite into new safe, effective drugs that have activity against all forms of the parasite, and which can be easily administered. Polyamines are essential metabolites that are required for cell growth. The polyamine biosynthetic enzymes, including ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (AdoMetDC) are essential to the parasite. a-difluoromethyornithine (DFMO), a suicide inhibitor of ODC, is the frontline treatment for late-stage T. gambiense, validating polyamine biosynthesis as a drug target in the parasite. Our group has systematically explored the potential for other enzymes in the polyamine biosynthetic pathway to be validated for the discovery of new anti-trypanosomal agents, leading to the validation of AdoMetDC as a highly promising target for HAT drug discovery. We demonstrated that AdoMetDC is essential to blood stage T. brucei parasites using genetic approaches and we discovered that the enzyme is regulated by a unique mechanism not found in mammalian cells. Starting from a previously identified human AdoMetDC inhibitor (MDL 73811) we identified new compounds (eg Genz 644131) that are potent irreversible inhibitors of T. brucei AdoMetDC, with nM activity against parasites in vitro, and that are curative in early stage mouse infection models. Genz 644131 however is not effective against late stage infection models in mice. Our objectives are to identify novel inhibitors of T. brucei AdoMetDC without this liability, and to undertake a lead optimization program around these inhibitors to identify a preclinical candidate with the potential to treat both species and both stages of HAT. We have developed a two-part strategy that should maximize the chances that we can achieve this goal. In Aim 1 we will undertake additional lead optimization work on the Genz 644131 series with the goal of identifying compounds with improved brain penetration and activity in the late stage model. In Aim 2 we will conduct a high throughput screen (HTS) to identify novel T. brucei AdoMetDC inhibitors. Compounds identified from the HTS will be validated, and two of the best series will be selected based on a target selection matrix that includes potency, selectivity, activity in whole cell assays and suitable ADME properties. An iterative lead optimization program of the top series will be prosecuted in Aim 3. In Aim 4 mechanism of action and resistance studies will be undertaken, providing supporting data on suitability of the identified candidates. DNDi has agreed to serve as program consultants on this project. Public Health Relevance: Human African sleeping sickness (HAT) is a fatal insect borne disease caused by a parasitic pathogen that leads to death if untreated. Current drug therapies have high toxicity and are difficult to administer in rural African settings where the disease is endemic. The goal of this proposal is to identify novel, safe and easily administered compounds that cure both early and late stage infections of this disease in animal models, resulting in the nomination of a preclinical development candidate for the treatment of HAT by the completion of the 5-year fund period.